Cell‐cell contact activation of fibroblasts increases the expression of matrix metalloproteinases

Figures & data

Table I. Expression profile of matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) as analyzed by microarrays.

	Description	Average signal (Present/Absent) ^a		Affymetrix ID	Fold increase in intensity
		Monolayers	Spheroids
MMP‐1	Matrix metalloproteinase 1 (interstitial collagenase)	3 784 (P)	22 117 (P)	204475_at	* 5.84 (P = 0.0435)
MMP‐2	Matrix metalloproteinase 2 (gelatinase A, 72 kDa gelatinase)	10 467 (P)	17 240 (P)	201069_at	1.65
MMP‐3	Matrix metalloproteinase 3 (stromelysin 1, progelatinase)	14 013 (P)	17 932 (P)	205828_at	1.28
MMP‐9	Matrix metalloproteinase 9 (gelatinase B, 92 kDa gelatinase)	470 (P)	1 484 (P)	203936_s_at	3.157
MMP‐10	Matrix metalloproteinase 10 (stromelysin 2)	114 (A)	12 077 (P)	205680_at	* 105.9 (P = 0.0493)
MMP‐11	Matrix metalloproteinase 11 (stromelysin 3)	114 (A)	404 (P)	203878_s_at	3.55
MMP‐12	Matrix metalloproteinase 12 (macrophage elastase)	152 (P)	163 (P)	204580_at	1.03
MMP‐14	Matrix metalloproteinase 14 (MT1‐MMP, membrane‐type)	230 (P)	1 292 (P)	217279_x_at	* 5.62 (P = 0.0489)
MMP‐19	Matrix metalloproteinase 19 (stromelysin 4)	431 (P)	860 (P)	204574_s_at	1.6
MMP‐24	Matrix metalloproteinase 24 (MT5‐MMP, membrane‐type)	255 (P)	296 (P)	49679_s_at	1.16
TIMP‐1	Tissue inhibitor of metalloproteinase 1	21 469 (P)	28 442 (P)	201666_at	1.33
TIMP‐2	Tissue inhibitor of metalloproteinase 2	1 151 (P)	648 (P)	203167_at	0.56
TIMP‐3	Tissue inhibitor of metalloproteinase 3	1 326 (P)	1 174 (P)	201149_s_at	0.89

Notes: ‘Fold increase in intensity’ indicates changes in expression intensity comparing spheroid cultures to corresponding monolayer cultures at 36 hours after seeding of cells. ‘Affymetrix ID’ refers to oligonucleotide numbering on the GeneChip Array U133A (Affymetrix). * indicates a significant difference (P<0.05) as compared to the corresponding monolayer. Transcripts for MMP‐7, MMP‐8, MMP‐13, MMP‐15, MMP‐16, MMP‐17, MMP‐20, MMP‐23, MMP‐25, MMP‐26, MMP‐27, MMP‐28 and TIMP‐4 were not expressed either in monolayer or spheroid cultures. ^a The signal is considered to be absent (A) or present (P) based on Affymetrix equation. Complete analysis of microarray data will be published elsewhere.

Figure 1. Immunoblots showing expression of(A) MMP‐1; (B) MMP‐10; (C) MT1‐MMP; and (D) TIMP‐1 in cell culture media of cell‐cell activated fibroblasts at the indicated time points. (E) Expression of MT1‐MMP in spheroid lysates at the indicated time points.

Figure 2. In situ hybridizations (ISH) of (A) uPA mRNA; (B) tPA mRNA; and (F) a sense, negative control for uPA mRNA, are made with Blue Map staining. (C) uPA immunostaining (IHC); (D) tPA IHC; and (E) CD‐68 IHC (negative control) are made with brown DAB staining. Time point was 72 h. Arrows show some positive signals of ISH. 40× objective was used in microscopy.